The effect of incubation time on Enzyme-Linked Immunosorbent Assay (ELISA) / Anisah Haji Musa

A typical Enzyme-Linked Immunosorbent Assay (ELISA) needs 2 to 3 hours to complete the test. The aim of this study is to analyze the effect of incubation time on ELISA. There are 3 incubation steps in ELISA procedures. Only 2 of the steps are involved in this study which are, the incubation for antibody in the serum react with anti-human IgM coated in microwells and incubation for antigen-Mab complex react with remaining antibody after washing procedure. Three incubation times which are 60 minutes, 45 minutes and 30 minutes were tested. The assay used E-DEN01M test kit with the manufacturers' protocol as the reference. Two strips of assays wells comprise of 11 samples, a negative control, a positive control, and triplicates calibrators were performed in each run. From the assay, the cut-off value (COV) was calculated. The results showed a significant difference of absorbance values for shorter incubation time. The COV reduced as the time was reduced. The test method with shorter incubation time cannot be applied as the results were invalid. The 60 minutes is the optimum incubation time for this ELISA.